This is a multi-investigator proposal to study a novel means of enhancing the mechanism by which retinoic acid (RA) causes growth arrest and differentiation by exploiting RA-induced expression and signaling by the CD38 receptor. RA is a cancer chemopreventive agent. It is a prohormone provided in the diet and is a common dietary insufficiency. RA can convert immature neoplastically transformed cells to a mature differentiated phenotype. The proposed studies utilize immature, uncommitted human leukemia granulocytic-monocytic precursor cells which undergo G0 arrest and either myeloid or monocytic differentiation when treated with RA or vitamin D3 (D3) respectively. RA causes activation of MAPK signaling in the process of inducing terminal myeloid differentiation/G0 arrest. Surveys of RA-induced gene expression from our and other laboratories revealed that CD38 is the earliest known receptor induced by RA. Crippling RA-induced CD38 expression cripples RA-induced differentiation, and ectopic over expression of CD38 enhances RA-induced differentiation. CD38 could potentially function through either ectoenzyme activity or receptor signaling to support differentiation: (1) The ectoenzyme activity catalyzes production of ADPR from NAD; and cADPR is able to mobilize calcium, a known regulator of differentiation and proliferation. (2) CD38 is also capable of dimerizing, which is another well known initiator of MAPK signaling. (3) CD38 has a cytosolic tail capable of binding to MAPK signaling molecules, in particular the c-Cbl adaptor. The proposed studies focus on the potential role of these functions in effecting the cellular outcome attributed to RA, namely differentiation and G0 cell cycle arrest. The proposed goals are to use small molecule probes that bind CD38, CD38 mutants, and X-ray crystallographic structures to characterize the mechanism by which CD38 promotes cell differentiation and arrest. The Specific Aims are to (1) determine the contribution of the CD38 ectoenzyme activity to RA-induced differentiation; (2) determine the contribution of receptor dimerization and the consequential cytosolic MAPK signaling complex; (3) determine the structure of the CD38-c-Cbl receptor-adaptor complex to elucidate the nature of the physical interfaces available for partners or intervention. In sum we intend to identify the CD38 activity needed to propel RA-induced differentiation. The Impact of these studies is a potential paradigm shift in understanding how RA works that will point to means of enhancing its mechanism of action. The Significance of the studies is that they could provide the basic mechanistic insight into a novel means of enhancing the chemopreventive, chemotherapeutic effects of RA. The Innovation is that a new paradigm based on receptor expression and signaling activities is proposed as a mechanism of action for RA in reverting neoplastic transformation. The basic Hypothesis is thus that RA-induces the early expression of CD38 which has different signal generating capabilities, and that these different capabilities could be exploited to enhance and control the differentiation/arrest program elicited by RA. The ultimate goal is to enhance the effects of RA.